Project Summary/Abstract The overarching goal of the parent grant has been to gain critical insights into the fundamentals of kinesin motor structure and function and to extrapolate this understanding to the inner workings of the cell. Kinesin superfamily members share a common catalytic domain yet participate in a wide range of cellular functions. Sequence differences modify the catalytic properties and microtubule interactions that are critical for the specific function of a kinesin. In this funding cycle our specific aims are directed to establish the mechanistic, mechanical, and structural features that are shared by Kinesin-2 KIF3AB and KIF3AC, and to define the mechanism by which heterodimerization modulates the distinct motile properties of KIF3AC in comparison to KIF3AB and other processive kinesins. The supplemental funds are requested specifically to purchase a GE Healthcare AKTA pure protein purification system which is essential for our mechanistic studies to define the presteady-state ATPase kinetics of the microtubule-based ATPase cycle. Our present AKTA system is 11 years old, and the model is due to be discontinued in the coming year which will result in a cost increase in the annual service contract followed by a gradual decrease in company support to no support in ~5 years. This supplement will ensure our ability to purify proteins in large amounts required to accomplish the specific aims as outlined in the parent grant.